International Publication WO 00/70067, published Nov. 23, 2000, is directed to a rice actin 2 promoter and actin 2 intron and methods for the use thereof. Environment or stress resistance to drought (see corresponding U.S. Pat. No. 6,429,357, at cols. 19 and 20) is described by introducing genes encoding for trehalose-6-phosphate synthase and through subsequent action of native phosphatases in the cell or by introduction and coexpression of a specific phosphatase resulting in trehalose which is a protective compound able to mitigate the effects of stress.
U.S. Pat. No. 5,925,804 is directed to the increase in the production of Trehalsoe in plants using an E. coli trehalose phosphate synthase gene, see cols. 7 and 8.
Seo H S at al., Appl Environ. Microbiol, 65:2484-2490, (2000), which relates to the characterization of a bifunctional fusion enzyme (TPSP) of trehalose-6-phosphate synthetase and trehalose-6-phosphate phophatase of Escherichia coli. 
Trehalose (α-D-glucopyranosyl-[1,1]-α-D-glucopyranose) is a non-reducing diglucoside and therefore does not react with amino acids or proteins as part of Maillard browning. Trehalose is found in various organisms, including bacteria, algae, fungi, yeast, insects and some plants, and serves not only as a carbohydrate reservoir but also as a protective agent against a variety of physical and chemical stresses (see, Elbein A, Adv. Carbohydr. Chem. Biochem., 30:227-256, 1974; Eleutherio E C A et al., Cryobiology, 30:591-596, 1993; Strom A R and Kaasen I, Mol. Microbiol., 8:205-210, 1993; van Laere A, FEMS Microbiol. Rev., 63:201-210, 1989; and Wiemken A, J. Gen, Microbiol., 58:209-217, 1990). Further, it has been known that trehalose shows a high water-retention activity under dry conditions to maintain the fluidity of the cell membranes and allow the plant to have a resistance against naturally occurring stresses during cycles of dehydration and rehydration (see, Leslie S B et al., Appl. Environ. Microbiol., 61:3592-3597, 1995; Drennan P M et al., J. Plant Physiol., 142:493-496, 1993; and Muller J et al., Plant Sci., 112:1-9, 1995). Such effect of trehalose on stress resistance has been demonstrated for cryptobiotic plant species such as S. leidophylla having resistance against dehydration. In this regard, it has been reported that trehalose accumulates to the level of 12% of plant dry weight during dehydration of such plant species, whereas trehalose accumulation is reduced during rehydration (see, Goddijn O J M and van Dun K, Trends Plant Sci., 4:315-319, 1999).
By virtue of such activity of trehalose, it has been attempted to increase stress resistance of plants. Up to the present, transgenic plants that express trehalose-6-phosphate synthetase (PTS) gene and/or trehalose-6-phosphate phosphatase (TPP) gene from E. coli or yeast in dicotyledon plants have been found. These transgenic plants express trehalose generally at a very low level. However, in these transgenic plants, although the stress resistance was somewhat increased, adverse effects appeared such as severe growth disturbance and warped roots. These adverse effects were exhibited even in the absence of trehalose accumulation (see, Holmstrom K-O et al., Nature, 379:683-684, 1996; Goddijn O J M et al., Plant Physiol, 113:181-1990, 1997; Muller et al., Plant Sci, 147:37-47, 1999; Pilon-Smits E A H et al., J. Plant Physiol., 152:525-532, 1998; and Romeo C et al., Planta, 201:293-297, 1997).
In the production of food for human welfare and existence, monocot plants, including rice, barley, wheat, maize, etc., are regarded as being commercially valuable plants. Therefore, a lot of effort has been exerted to increase the productivity and quality of such crops. Particularly, continuous efforts have been made in order to produce crops having resistance against abiotic natural conditions, such as drought, an increase in salt concentration, low temperature, etc.
The explosive increase in world population, along with the continuing deterioration of arable land, scarcity of fresh water, and increasing environmental stress pose serious threats to global agricultural production and food security. Despite focused efforts to improve major crops for resistance to abiotic stresses such as drought, excessive salinity, and low temperature by traditional breeding, success has been limited (Boyer, J. S., “Plant Productivity and Environment,” Science, 218:443-448 (1982)). This lack of desirable progress is attributable to the fact that tolerance to abiotic stress is a complex trait that is influenced by coordinated and differential expression of a network of genes. Fortunately, it is now possible to use transgenic approaches to improve abiotic stress tolerance in agriculturally important crops with far fewer target traits than had been anticipated (Zhang et al., “Engineering Salt-Tolerant Brassica Plants: Characterization of Yield and Seed Oil Quality in Transgenic Plants with Increased Vacuolar Sodium Accumulation,” Proc. Natl. Acad. Sci. USA, 98:12832-12836 (2001)).
Abiotic stresses can directly or indirectly affect the physiological status of an organism by altering its metabolism, growth, and development. A common response of organisms to drought, salinity, and low-temperature stresses is the accumulation of sugars and other compatible solutes (Hare et al., “Dissecting the Roles of Osmolyte Accumulation During Stress,” Plant Cell Environ., 21:535-553 (1998)). These compounds serve as osmoprotectants and, in some cases, stabilize biomolecules under stress conditions (Hare et al., “Dissecting the Roles of Osmolyte Accumulation During Stress,” Plant Cell Environ., 21:535-553 (1998); Yancey et al., “Living with Water Stress: Evolution of Osmolyte Systems,” Science, 217:1214-1222 (1982)). One such compound is trehalose, a nonreducing disaccharideof glucose, which plays an important physiological role as an abiotic stress protectant in a large number of organisms, including bacteria, yeast, and invertebrates (Crowe et al., “Anhydrobiosis,” Annu. Rev. Physiol., 54:579-599 (1992)). Trehalose has been shown to stabilize dehydrated enzymes, proteins, and lipid membranes efficiently, as well as protect biological structures from damage during desiccation. In the plant kingdom, most species do not seem to accumulate detectable amounts of trehalose, with the notable exception of the highly desiccation-tolerant “resurrection plants” (Wingler, “The Function of Trehalose Biosynthesis in Plants,” Phytochemistry, 60:437-440 (2002)). The recent discovery of homologous genes for trehalose biosynthesis in Selaginella lepidophylla, Arabidopsis thaliana, and several crop plants suggests that the ability to synthesize trehalose may be widely distributed in the plant kingdom (Goddijn et al., “Trehalose Metabolism in Plants,” Trends Plant Sci., 4:315-319 (1999)). A putative plant gene for trehalose-6-phosphate synthase (TPS) can complement a Δtps1 mutant yeast strain, suggesting that the plant and yeast gene products are functionally similar (Zentella et al., “A Selaginella lepidophylla Trehalose-6-Phosphate Synthase Complements Growth and Stress-Tolerance Defects in a Yeast tps1 Mutant,” Plant Physiol., 119:1473-1482 (1999)).
In bacteria and yeast, trehalose is synthesized in a two-step process: trehalose-6-phosphate is first formed from UDP-glucose and glucose-6-phosphate in a reaction catalyzed by TPS. Trehalose-6-phosphate is then converted to trehalose by trehalose-6-phosphate phosphatase (TPP) (Goddijn et al., “Trehalose Metabolism in Plants,” Trends Plant Sci., 4:315-319 (1999)). Metabolic engineering for enhanced accumulation of trehalose in plants has been the recent focus of attention in some model dicot plants (Holmstrom et al., “Drought Tolerance in Tobacco,” Nature, 379:683-684 (1996); Goddijn et al., “Inhibition of Trehalase Activity Enhances Trehalose Accumulation in Transgenic Plants,” Plant Physiol., 113:181-190 (1997); Romero et al., “Expression of the Yeast Trehalose-6-Phosphate Synthase Gene in Transgenic Tobacco Plants: Pleiotropic Phenotypes Include Drought Tolerance,” Planta, 201:293-297 (1997); Pilon-Smits et al., “Trehalose-Producing Transgenic Tobacco Plants Show Improved Growth Performance Under Drought Stress,” J. Plant Physiol., 152:525-532 (1998)). However, in these previous studies, constitutive overexpression of TPS and/or TPP genes from yeast or Escherichia coli in tobacco or potato plants resulted in undesirable pleiotropic effects, including stunted growth and altered metabolism under normal growth conditions (Goddijn et al., “Inhibition of Trehalase Activity Enhances Trehalose Accumulation in Transgenic Plants,” Plant Physiol., 113:181-190 (1997); Romero et al., “Expression of the Yeast Trehalose-6-Phosphate Synthase Gene in Transgenic Tobacco Plants: Pleiotropic Phenotypes Include Drought Tolerance,” Planta, 201:293-297 (1997); Pilon-Smits et al., “Trehalose-Producing Transgenic Tobacco Plants Show Improved Growth Performance Under Drought Stress,” J. Plant Physiol., 152:525-532 (1998)).
The present invention is directed, inter alia, to producing transgenic monocot plants with improved low temperature stress, water stress, and salt stress tolerance.